Because of its critical role in Myxococcus xanthus fruiting body formation and its potential role as a genetic switch, the devRS operon will be further analyzed, emphasizing the negative autoregulation. The autoregulatory protein will first be identified by making deletions in the genes identified and determining which one affects the accumulation of RNA. Then, it will be determined if the regulation is at the level of RNA stability or at the level of repression of transcription. The RNA stability will be examined by determining the half-life of the RNA in the wild type and mutant strains. If it is not different in these strains, then repression of transcription will be examined. The promoter region will first be sequenced and the transcriptional start site will be determined using S1 mapping and primer extension. Then a mutational analysis of the promoter region will be performed to determine sites required for repression. After this, the autoregulatory protein will be purified using current fusion protein technology and interactions with the RNA or DNA will be analyzed.
